DNA Replication Initiation Is Blocked by a Distant Chromosome-Membrane Attachment.
DNA Replication Initiation Is Blocked by a Distant Chromosome-Membrane Attachment.
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DOI：
10.1016/j.cub.2015.06.058
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发表时间：
2015-08-17
期刊：
影响因子：
--
通讯作者：
Bates D
中科院分区：
文献类型：
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作者：
Magnan D;Joshi MC;Barker AK;Visser BJ;Bates D
虽然几十年来人们已经认识到染色体结构调节了复制起始点的启动能力，但对于细胞如何或是否主动调节结构来直接启动却知之甚少。我们报道了大肠杆菌细胞中染色体和细胞膜之间的局部诱导蛋白系链对复制起始具有快速和完全的阻断作用。系链由跨膜和转录抑制融合蛋白与一系列操作序列结合组成，可以放置在距离起源1兆碱基处而不会损失外显子。系绳诱导的起始阻断对预先存在的复制叉的延伸没有影响，也不会引起细胞或DNA损伤。拴系细胞的全基因组和位点特异性荧光DNA标记表明，整体的类核结构和染色体组织被破坏。通过RNA测序分析的基因表达模式表明，系缚诱导了全局超螺旋变化，这可能与复制起始不相容。讨论了系绳诱导的起始阻断和利福平治疗之间的相似之处，以及染色体结构中程序化变化在复制控制中的作用。
Although it has been recognized for several decades that chromosome structure regulates the capacity of replication origins to initiate, very little is known about how or if cells actively regulate structure to direct initiation. We report that a localized inducible protein tether between the chromosome and cell membrane in E. coli cells imparts a rapid and complete block to replication initiation. Tethers, composed of a trans-membrane and transcription repressor fusion protein bound to an array of operator sequences, can be placed up to one megabase from the origin with no loss of penetrance. Tether-induced initiation blocking has no effect on elongation at pre-existing replication forks and does not cause cell or DNA damage. Whole-genome and site-specific fluorescent DNA labeling in tethered cells indicates that global nucleoid structure and chromosome organization are disrupted. Gene expression patterns, assayed by RNA sequencing shows that tethering induces global supercoiling changes, which are likely incompatible with replication initiation. Parallels between tether-induced initiation blocking and rifampicin treatment, and the role of programmed changes in chromosome structure in replication control are discussed.